Targeted Deletion of the gldB Gene to Disrupt Gliding Motility in Cellulophaga lytica

Presenter: Mehul Puri

Faculty Sponsor: Milana Vasudev

School: UMass Dartmouth

Research Area: Biomedical Engineering

Session: Poster Session 3, 1:15 PM - 2:00 PM, Auditorium, A21

ABSTRACT

The rod-shaped gram-negative bacteria Cellulophaga Lytica grows in marine ecosystems and has the unique ability of producing iridescence. Iridescence, commonly found in eukaryotes such as birds and fishes, is uncommon to bacteria. C. lytica produces this glitter-like coloration due to the interactions between light and the bacteria’s physical structures arranged in a periodic geometry. Interestingly, the bacteria’s biofilms produce an intense iridescence without a flagella or pili on the cells. Instead, cells depend on gliding motility to transport themselves and form biofilms. Sequencing of C. lytica's genome revealed that the gldB gene, which produces the gld protein, is responsible for the bacteria's gliding motility.​

The suicide vector pYT313 is an integrative plasmid that cannot replicate in the target host C. lytica. Its unique design lacks an origin of replication in non-E. coli species, allowing it to replace the target gliding motitlity gene (gldB) with the erythromycin resistance gene (ermF). ​

In this study, the pYT313 suicide vector is modified to contain the ermF_KO insert such that it can integrate into C. lytica, knockout the gldB gene, and insert erythromycin resistance gene.​ To do this, we need to modify the pYT313 suicide vector for gene deletion. Golden gate assembly is conducted to ligate the ermF_KO insert into pYT313. Conjugation between modified pYT313 transformed E. coli and C. lytica will allow for direct DNA transfer through a two-step allelic exchange. ​This will allow us to draw a correlation between the gliding motility of C. lytica and its iridescence through gene deletion​

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